Mammalian cell lines are used to study the cellular processes that regulate genome stability and to determine how those processes can be perturbed by specific gene mutations and/or by environmental mutagens/carcinogens. Our studies use both rodent and human cells. We have developed primary rat and mouse cell strains from the transgenic Big Blue? Fischer-344 rat and B6C3F1 mouse. The intent of these studies was to define normal (i.e., nontransformed) cell strains derived from the Big Blue? rat or mouse for use in studies designed to complement the evaluation of a drug or chemical induced mutagenesis and/or clastogenic activity in vivo. We were able to demonstrate that these primary mouse (BBM1) and rat (BBR1) cells are genetically stable and mostly diploid. Both cell strains have low spontaneous frequencies of mutation at the lacI and cII loci as well as low frequencies of sister chromatid exchange and micronuclei formation which makes them useful model systems for molecular toxicology studies. Using Big Blue? Rat2 cells we demonstrated the utility and defined parameters for using the bacteriophage lambda cII locus for mutation studies, thereby provideing a convenient alternative to lacI. In addition, we have continued our efforts in evaluating the mutagenic consequnce of defects in human mismatch repair (MMR) genes following treatment with DNA damaging agents. Genomic stability in cell lines is known to be affected by defects in any one of the five known human MMR genes. The specific genes implicated in the human MMR pathway include homologues of the bacterial MMR proteins MutS and MutL, including MSH2, MSH3, MSH6 (GTBP), MLH1 and PMS2. We have previously shown that cytotoxicity to 6-thioguanine and to methylmethane sulfonate (MMS) is mediated through MMR, as indicated by an increased resistance to either 6-TG or MMS in MMR-deficient cells. In the absence of MMR processing of the damage induced by these agents mutations result. This result was surprising for MMS since the primary lesion results in the rapid formation of an abasic site. The implication of these data is that MMR may be involved in the repair of abasic sites. Indeed, the sequence spectra of mutations arising at the HPRT locus following MMS treatment of MMR-deficient and MMR-proficient cells strongly suggest that the sites and types of mutations observed result from the absence of repair of abasic sites.